The influence of non-canonical structures on the p53 isoforms binding to DNA

abstrakt

angličtina

Protein p53 is one of the most studied tumor suppressor protein plays important roles in regulating basic biological processes including cell cycle, apoptosis, senescence and metabolism. The human p53 gene contains alternative promoters and thank to alternative splicing could be expressed in several isoforms. P53 protein function is realized by binding to specific DNA response elements (RE) and transactivation of target genes. Here we presented results of p53alpha, p53beta and p53gamma isoforms DNA interaction studied by yeast isogenic system for in vivo transactivation studies in chromosomal structures to protein-DNA binding. WE used a panel of Saccharomyces cerevisiae haploid isogenic strains, except for the different p53 target site with propensity to form different DNA structures located upstream of the luciferase reporter gene. The targeting of p53 target sequence of interest by the replacement of the ICORE cassette, using transfected single strand oligonucleotides, was performed following the Delitto Perfetto technique. Yeast isogenic strains differing in the p53 target site (with and without propensity to form cruciform structure were transformed with plasmid for the expression of p53alpha isoform protein. Our results show that transactivation is in vivo correlated better with the relative propensity of a RE to form cruciform structure that to its expected in theoretical DNA binding affinity DNA binding affinity. These results point to the fact that structural features of DNA are an important determinant of its DNA-binding and transactivation function.

p53, inverted repeats, protein-DNA binding

PORUBIAKOVÁ, O.; BOHÁLOVÁ, N.; BRÁZDA, V.

27. 11. 2020

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